1. Field of the Invention
This invention relates generally to a process of rendering an allograft or xenograft ("foreign graft") nonthrombogenic and substantially nonimmunogenic for transplantation purposes. More particularly, the process involves creating an ex vivo interface between donor and recipient by coating the vascular endothelial cells, lining the lumen of blood vessels within the vascular network of the graft, with endothelial extracellular matrix immunologically tolerable (i.e., tolerated as an autograft) to the recipient; thereby providing a substantially nonimmunogenic, nonthrombogenic interface which provides a surface that enhances the re-endothelialization of the graft with host/recipient endothelium in "immunomodifying" the graft.
2. Description of the Background and Related Art
Organ transplantation is the therapy of choice for endstage organ failure. In the case of kidneys, transplantation provides for increased life expectancy, enhanced quality of life, and is more cost-effective than maintaining patients on hemodialysis. In the case of extrarenal organs, transplantation is life-saving since no equivalent to hemodialysis exists for these organs. The limiting factor today in the number of transplant procedures performed is the severe shortage of organs (Annual Report for the U.S. Scientific Registry for Organ Transplantation and the Organ Procurement and Transplantation Network, 1990).
To address organ shortage problems by providing ways in which organs/grafts can be better preserved for transplantation, a warm preservation technology (between 18.degree. C. to 35.degree. C.) was developed with others by the present inventors. U.S. patent application Ser. No. 08/029,459 discloses a method of warm preservation of a tissue intended for transplantation using a preservation solution that may be used for the initial flushing, and as a perfusate for storage of the tissue/organ (the disclosure of which is herein incorporated by reference). U.S. patent application Ser. No. 08/033,629 now abandoned discloses a method of warm preservation of a tissue intended for transplantation using a perfusate, supplemented with a perfluorochemical emulsion, for storage of the tissue/organ (the disclosure of which is herein incorporated by reference).
However, other problems need be addressed in the transplantation process. For example, without immunosuppressive agents/drugs, the onset of rejection of an anallograft may occur approximately seven days posttransplant. In the case of a xenograft, rejection may occur within a matter of minutes or hours. Rejection of the transplanted graft is an immunological assault in which the recipient's immune system recognizes the graft as "foreign" and attempts to eliminate the transplanted graft. The current theory of rejection in transplantation involves both the humoral and cell-mediated immune responses. The humoral response involves binding of the recipient's naturally occurring antibodies to the donor's vascular endothelial cells lining the blood vessels within the transplanted graft. Antibody deposition leads to the activation of the complement cascade which mediates a cytotoxic phenomenon which can directly damage or kill the endothelial cells. In addition the complement cascade leads to the activation of the endothelial cells which causes subsequent change in the anticoagulant environment. More specifically, the vascular endothelium normally provides a nonthrombogenic surface; therefore, when activated by the immune system during the rejection process, the endothelial lining transforms into a procoagulant environment. The resultant prothrombotic (thrombogenic) surfaces then attract polymorphonuclear cells and platelets, resulting in the endothelium being damaged and causing separation from the underlying substratum, and ultimately, severe thrombosis of the graft. The cell-mediated response is thought to involve not only the T-cell cytotoxicity-effector mechanisms, but also NK and K cell activity.
The standard approach to mitigate the rejection process is to treat the transplant recipient daily with an immunosuppressive regimen. However, currently immunosuppressive regimens are systemic; i.e., in addition to suppressing immune function against the transplanted graft, immune function which protects the recipient from other processes (such as infections) is suppressed. Further, the currently available immunosuppressives may cause substantial non-specific, toxic effects on cell types besides cells comprising the immune system.
For example, a typical immunosuppressive regimen comprises four therapeutic components--steroids, azathioprine, cyclosporin, and anti-T cell antibodies. The use of such therapy may lead to systemic secondary complications. Side effects of long term steroid use include bone disease, cataracts, diabetes, ulcers, pancreatitis and perforation of the colon. The use of azathioprine can cause hematologic toxicity resulting in leukopenia and/or thrombocytopenia; and can cause gastrointestinal complications. Additionally, use of azathioprine results in a significant risk of secondary infection and/or neoplasia. Cyclosporin is nephrotoxic, hepatotoxic, and results in a significant risk for development of hypertension, gingivitis, and neurologic tremors. Administration of anti-T cell antibodies can lead to cytokine-release syndrome, with the most serious complication being acute pulmonary edema. Gastrointestinal complications, typically with severe diarrhea, are common. Central nervous system symptoms may mimic aseptic meningitis, or cause seizures and encephalopathy. In addition to the serious complications with immunosuppressive agents, there is a need to improve efficacy of immunosuppressive therapy. Even in clinical transplantation with the greatly expanded arsenal of immunosuppressive agents, allograft rejection occurs approximately 25% of the time.
In attempting to solve problems associated with xenotransplantation, efforts are directed to mitigating the effect of natural occurring antibodies and the complement cascade. Two research groups (J. P. Richardson of Washington University and J. L. Platt of Duke University in collaboration with Nextran) have focused on temporarily removing the natural occurring antibody by plasmapheresis (or transfecting cells with human genes) with resultant blocking of the complement cascade. Plasmapheresis treatment is only temporary in that the antibody titer quickly returns when the treatments are discontinued. When the natural antibodies return in the circulation, the vascular rejection of the transplanted graft also occurs.
To avoid the systemic complications associated with systemic immunosuppressive therapy, and thereby leave the recipient's immune system intact, the development of a graft-specific therapy which is immunosuppressive or which shields or cloaks the transplanted graft from recipient's immune system is desirable. For example, one approach is to genetically alter a foreign graft (in this case, a xenograft) in the donor before xenotransplantation (Biomedical Business International Report, supra). Although several approaches have been taken, the most notable involves the incorporation of membrane associated inhibitors of the complement cascade into graft endothelial cells. There are a number of molecules which have been identified for their ability to inhibit the complement cascade at various points. To name a few, these molecules include: decay accelerating factor, CD46, homologous restriction factor and CDS9. Since these membrane associated inhibitors of complement are species-specific, it is hoped that membrane expression of the molecules in a graft will prevent the transplanted graft from being rejected. However, because the graft rejection process is multifaceted, involving several arms of the immune system, a number of genes would need to be incorporated into graft endothelial cells in order for the transplanted graft to be tolerated by the recipient.
U.S. Pat. No. 5,192,312 (C. Orton) discloses that cells of a graft cannot be masked to reduce immunogenicity. Rather, native cells of the graft must be removed, and replaced with allogeneic or autologous cells in a process involving incubation of the graft with growth factor and allogeneic or autologous fibroblasts prior to transplantation of the graft.
Another approach involves microencapsulated exocrine cells and hepatocytes in attempts to develop a synthetic pancreas and liver, respectively (Biomedical Business International Report, supra). Some form of cell attachment substrate is used to enhance function of the islet cells and hepatocytes. However, different substrata matrix lead to different morphology in the cultured cells. Collagen matrix grown hepatocytes had distinct morphology when compared to hepatocytes grown in MATRIGEL.TM.--an extracellular matrix secreted by a Engelberth-Holm-Swarm (EHS) tumor. The matrix components being used as substructure for cell attachment include collagen, fibronectin, laminin, and vitronectin (Biomedical Business International Report, supra). MATRIGEL.TM. is a reconstituted basement membrane from solubilized extract of the basement membrane from the EHS transplantable mouse tumor. Therefore, it is a mouse basement membrane derived from a transformed cell line. In as much, the matrix contains by-products of a transformed cell line such as angiogenic factors. While the EHS tumor-derived extracellular matrix contains laminin, type IV collagen, and heparin sulfate proteoglycans, the ratio of these components are distinct and are not representative of normal basement membranes (Kleinman et al., 1982, Biochemistry 22:4969). MATRIGEL.TM. and type I collagen have been used to support hepatocytes for a synthetic liver.
Thus, there exists a need for expanding the organ donor pool, and a corresponding need for a method to treat a foreign graft in such a way as to be tolerated in a recipient, i.e. thereby avoiding rejection by avoiding activation of a recipient's humoral and cell-mediated immune response to the transplanted graft. The method should also be graft-specific to avoid complications such as those associated with systemic immunosuppressive therapy.